PROJECT ABSTRACT The proposed study, Predictive biomarkers for the onset of immune-related adverse events associated with PD-1 blockade, addresses NCI?s Provocative Question 8 (PQ8): What are the predictive biomarkers for the onset of immune-related adverse events associated with checkpoint inhibition, and are they related to markers for efficacy? Programmed cell death-1 (PD-1) is an essential inhibitory receptor in T cells. Antibodies that block PD-1 augment anti-tumor immune responses and represent a powerful therapeutic paradigm. There are currently over 400 clinical trials targeting PD-1, highlighting its emerging potential in cancer immunotherapy. Unfortunately, 1 in 4 patients who receive anti-PD-1 treatment develop immune-related adverse events (irAEs), including autoimmunity. As such, elucidating the mechanisms of immune-tolerance breakdown, which lead to excessive inflammation in cancer patients treated with immune-checkpoint inhibitors (such as anti-PD-1), presents a formidable opportunity to understand the molecular underpinnings of irAEs and to identify predictive biomarkers of irAEs in order to develop safer next-generation immunotherapies. To achieve this goal it is essential to understand how PD-1 blockade reshapes homeostatic T cell responses. We propose to uncover the repertoire diversity and the specific gene expression signatures of antigen-experienced T cells from patients undergoing anti-PD-1 therapy, via two Specific Aims: 1) to define T cell-specific gene expression signatures that distinguish patients who develop irAEs following anti-PD-1 therapy; and 2) to test the hypothesis that anti-PD-1 therapy decreases T cell repertoire diversity in patients who develop irAEs. We anticipate that our findings will shed light on the molecular complexity of both T cell repertoire transition and signaling dynamics during the development of irAEs. Supported by preliminary data, this project will apply new approaches to uncover the molecular mechanisms underlying irAEs. The proposed project will combine transcriptional and clonal repertoire analyses of T cells from lung cancer patients treated with anti-PD-1, to elucidate mechanisms of self-tolerance breakdown in immunotherapies. Within the proposed two years, we will examine PD-1 associated signaling clusters pathways in the setting of irAEs. These studies will reveal the immuno-pathogenesis of irAEs and will likely reveal both predictive and severity biomarkers for irAEs.